Inbred corn line MN5

ABSTRACT

An inbred corn line, designated MN5, is disclosed. The invention relates to the seeds of inbred corn line MN5, to the plants and plant parts of inbred corn line MN5 and to methods for producing a corn plant, either inbred or hybrid, by crossing the inbred line MN5 with itself or another corn line. The invention further relates to methods for producing a corn plant containing in its genetic material one or more transgenes and to the transgenic plants produced by that method and to methods for producing other inbred corn lines derived from the inbred MN5.

BACKGROUND OF THE INVENTION

The present invention relates to a new and distinctive corn inbred line,designated MN5. All publications cited in this application are hereinincorporated by reference.

There are numerous steps in the development of any novel, desirableplant germplasm. Plant breeding begins with the analysis and definitionof problems and weaknesses of the current germplasm, the establishmentof program goals, and the definition of specific breeding objectives.The next step is selection of germplasm that possess the traits to meetthe program goals. The goal is to combine in a single variety or hybridan improved combination of desirable traits from the parental germplasm.These important traits may include higher yield, resistance to diseasesand insects, better stalks and roots, tolerance to drought and heat,reduction of grain moisture at harvest as well as better agronomicquality. With mechanical harvesting of many crops, uniformity of plantcharacteristics such as germination and stand establishment, growthrate, maturity and plant and ear height is important.

Choice of breeding or selection methods depends on the mode of plantreproduction, the heritability of the trait(s) being improved, and thetype of cultivar used commercially (e.g., F₁ hybrid cultivar, purelinecultivar, etc.). For highly heritable traits, a choice of superiorindividual plants evaluated at a single location will be effective,whereas for traits with low heritability, selection should be based onmean values obtained from replicated evaluations of families of relatedplants. Popular selection methods commonly include pedigree selection,modified pedigree selection, mass selection, recurrent selection, andbackcross breeding.

The complexity of inheritance influences choice of breeding method.Backcross breeding is used to transfer one or a few favorable genes fora heritable trait into a desirable cultivar. This approach has been usedextensively for breeding disease-resistant cultivars; nevertheless, itis also suitable for the adjustment and selection of morphologicalcharacters, color characteristics and simply inherited quantitativecharacters such as earliness, plant height or seed size and shape.Various recurrent selection techniques are used to improvequantitatively inherited traits controlled by numerous genes. The use ofrecurrent selection in self-pollinating crops depends on the ease ofpollination, the frequency of successful hybrids from each pollination,and the number of hybrid offspring from each successful cross.

Each breeding program should include a periodic, objective evaluation ofthe efficiency of the breeding procedure. Evaluation criteria varydepending on the goal and objectives, but should include gain fromselection per year based on comparisons to an appropriate standard,overall value of the advanced breeding lines, and number of successfulcultivars produced per unit of input (e.g., per year, per dollarexpended, etc.).

Promising advanced breeding lines are thoroughly tested per se and inhybrid combination and compared to appropriate standards in environmentsrepresentative of the commercial target area(s) for three or more years.The best lines are candidates for use as parents in new commercialcultivars; those still deficient in a few traits may be used as parentsto produce new populations for further selection.

These processes, which lead to the final step of marketing anddistribution, usually take from eight to twelve years from the time thefirst cross is made. Therefore, development of new cultivars is atime-consuming process that requires precise forward planning, efficientuse of resources, and a focus on clear objectives.

A most difficult task is the identification of individuals that aregenetically superior, because for most traits the true genotypic valueis masked by other confounding plant traits or environmental factors.One method of identifying a superior plant is to observe its performancerelative to other experimental plants and to a widely grown standardcultivar. If a single observation is inconclusive, replicatedobservations provide a better estimate of its genetic worth.

The goal of corn breeding is to develop new, unique and superior corninbred lines and hybrids. The breeder initially selects and crosses twoor more parental lines, followed by repeated self pollination or selfingand selection, producing many new genetic combinations. The breeder cantheoretically generate billions of different genetic combinations viacrossing, selfing and mutations.

Each year, the plant breeder selects the germplasm to advance to thenext generation. This germplasm is grown under unique and differentgeographical, climatic and soil conditions, and further selections arethen made, during and at the end of the growing season. The inbred lineswhich are developed are unpredictable. This unpredictability is becausethe breeder's selection occurs in unique environments, with no controlat the DNA level (using conventional breeding procedures), and withmillions of different possible genetic combinations being generated. Abreeder of ordinary skill in the art cannot predict the final resultinglines he develops, except possibly in a very gross and general fashion.This unpredictability results in the expenditure of large research fundsto develop a superior new corn inbred line.

The development of commercial corn hybrids requires the development ofhomozygous inbred lines, the crossing of these lines, and the evaluationof the crosses. Pedigree breeding and recurrent selection breedingmethods are used to develop inbred lines from breeding populations.Breeding programs combine desirable traits from two or more inbred linesor various broad-based sources into breeding pools from which inbredlines are developed by selfing and selection of desired phenotypes. Thenew inbreds are crossed with other inbred lines and the hybrids fromthese crosses are evaluated to determine which have commercialpotential.

Pedigree breeding is used commonly for the improvement ofself-pollinating crops or inbred lines of cross-pollinating crops. Twoparents which possess favorable, complementary traits are crossed toproduce an F₁. An F₂ population is produced by selfing one or severalF₁s or by intercrossing two F₁s (sib mating). Selection of the bestindividuals is usually begun in the F₂ population; then, beginning inthe F₃, the best individuals in the best families are selected.Replicated testing of families, or hybrid combinations involvingindividuals of these families, often follows in the F₄ generation toimprove the effectiveness of selection for traits with low heritability.At an advanced stage of inbreeding (i.e., F₆ and F₇), the best lines ormixtures of phenotypically similar lines are tested for potentialrelease as new cultivars.

Mass and recurrent selections can be used to improve populations ofeither self- or cross-pollinating crops. A genetically variablepopulation of heterozygous individuals is either identified or createdby intercrossing several different parents. The best plants are selectedbased on individual superiority, outstanding progeny, or excellentcombining ability. The selected plants are intercrossed to produce a newpopulation in which further cycles of selection are continued.

Backcross breeding has been used to transfer genes for a simplyinherited, highly heritable trait into a desirable cultivar or inbredline which is the recurrent parent. The source of the trait to betransferred is called the donor parent. The resulting plant is expectedto have the attributes of the recurrent parent (e.g., cultivar) and thedesirable trait transferred from the donor parent. After the initialcross, individuals possessing the phenotype of the donor parent areselected and repeatedly crossed (backcrossed) to the recurrent parent.The resulting plant is expected to have the attributes of the recurrentparent (e.g., cultivar) and the desirable trait transferred from thedonor parent.

Descriptions of other breeding methods that are commonly used fordifferent traits and crops can be found in one of several referencebooks (e.g., Allard, Principles of Plant Breeding, John Wiley and Son,pp. 115-161, 1960, Fehr, 1987).

Proper testing should detect any major faults and establish the level ofsuperiority or improvement over current cultivars. In addition toshowing superior performance, there must be a demand for a new cultivarthat is compatible with industry standards or which creates a newmarket. The introduction of a new cultivar will incur additional coststo the seed producer, the grower, processor and consumer for specialadvertising and marketing, altered seed and commercial productionpractices, and new product utilization. The testing preceding release ofa new cultivar should take into consideration research and developmentcosts as well as technical superiority of the final cultivar. Forseed-propagated cultivars, it must be feasible to produce seed easilyand economically.

Once the inbreds that give the best hybrid performance have beenidentified, the hybrid seed can be reproduced indefinitely as long asthe homogeneity of the inbred parent is maintained. A single-crosshybrid is produced when two inbred lines are crossed to produce the F₁progeny. A double-cross hybrid is produced from four inbred linescrossed in pairs (A×B and C×D) and then the two F₁ hybrids are crossedagain (A×B)×(C×D). Much of the hybrid vigor exhibited by F₁ hybrids islost in the next generation (F₂). Consequently, seed from hybridvarieties is not used for planting stock.

Hybrid corn seed is typically produced by a male sterility system or byincorporating manual or mechanical detasseling. Alternate strips of twocorn inbreds are planted in a field, and the pollen-bearing tassels areremoved from one of the inbreds (female). Providing that there issufficient isolation from sources of foreign corn pollen, the ears ofthe detasseled inbred will be fertilized only from the other inbred(male), and the resulting seed is therefore hybrid and will form hybridplants.

The laborious, and occasionally unreliable, detasseling process can beavoided by using cytoplasmic male-sterile (CMS) inbreds. Plants of a CMSinbred are male sterile as a result of factors resulting from thecytoplasmic, as opposed to the nuclear, genome. Thus, thischaracteristic is inherited exclusively through the female parent incorn plants, since only the female provides cytoplasm to the fertilizedseed. CMS plants are fertilized with pollen from another inbred that isnot male-sterile. Pollen from the second inbred may or may notcontribute genes that make the hybrid plants male-fertile. Seed fromdetasseled fertile corn and CMS produced seed of the same hybrid can beblended to insure that adequate pollen loads are available forfertilization when the hybrid plants are grown.

There are several methods of conferring genetic male sterilityavailable, such as multiple mutant genes at separate locations withinthe genome that confer male sterility, as disclosed in U.S. Pat. Nos.4,654,465 and 4,727,219 to Brar et al. These and all patents referred toare incorporated by reference. In addition to these methods, Albertsenet al., U.S. Pat. No. 5,432,068, have developed a system of nuclear malesterility which includes: identifying a gene which is critical to malefertility, silencing this native gene which is critical to malefertility; removing the native promoter from the essential malefertility gene and replacing it with an inducible promoter; insertingthis genetically engineered gene back into the plant; and thus creatinga plant that is male sterile because the inducible promoter is not “on”resulting in the male fertility gene not being transcribed. Fertility isrestored by inducing, or turning “on”, the promoter, which in turnallows the gene that confers male fertility to be transcribed.

There are many other methods of conferring genetic male sterility in theart, each with its own benefits and drawbacks. These methods use avariety of approaches such as delivering into the plant a gene encodinga cytotoxic substance associated with a male tissue specific promoter oran anti-sense system in which a gene critical to fertility is identifiedand an antisense to that gene is inserted in the plant (see,Fabinjanski, et al. EPO 89/0301053.8 publication no. 329,308 and PCTapplication PCT/CA90/00037 published as WO 90/08828).

Another version useful in controlling male sterility makes use ofgametocides. Gametocides are not a genetic system, but rather a topicalapplication of chemicals. These chemicals affect cells that are criticalto male fertility. The application of these chemicals affects fertilityin the plants only for the growing season in which the gametocide isapplied (see Carlson, G. R., U.S. Pat. No. 4,936,904). Application ofthe gametocide, timing of the application, and genotype often limit theusefulness of the approach.

Corn is an important and valuable field crop. Thus, a continuing goal ofplant breeders is to develop stable, high yielding corn hybrids that areagronomically sound. The reasons for this goal are obviously to maximizethe amount of ears or kernels produced on the land used and to supplyfood for both humans and animals. To accomplish this goal, the cornbreeder must select and develop corn plants that have the traits thatresult in superior parental lines for producing hybrids.

The foregoing examples of the related art and limitations relatedtherewith are intended to be illustrative and not exclusive. Otherlimitations of the related art will become apparent to those of skill inthe art upon a reading of the specification.

SUMMARY OF THE INVENTION

The following embodiments and aspects thereof are described inconjunction with systems, tools and methods which are meant to beexemplary and illustrative, not limiting in scope. In variousembodiments, one or more of the above-described problems have beenreduced or eliminated, while other embodiments are directed to otherimprovements.

According to the invention, there is provided an inbred corn line,designated MN5. This invention thus relates to the seeds of inbred cornline MN5, to the plants or parts thereof of inbred corn line MN5, toplants or parts thereof having all the physiological and morphologicalcharacteristics of inbred corn line MN5 and to plants or parts thereofhaving all the physiological and morphological characteristics of inbredcorn line MN5 listed in Table 1. Parts of the inbred corn plant of thepresent invention are also provided, such as e.g., pollen obtained froman inbred plant and an ovule of the inbred plant.

In another aspect, the present invention provides regenerable cells foruse in tissue culture of inbred corn plant MN5. The tissue culture willpreferably be capable of regenerating plants having all thephysiological and morphological characteristics of the foregoing inbredcorn plant. Preferably, the cells of such tissue cultures will beembryos, meristematic cells, seeds, callus, pollen, leaves, anthers,roots, root tips, silk, flowers, kernels, ears, cobs, husks, stalks orthe like. Protoplasts produced from such tissue culture are alsoincluded in the present invention. The corn plants regenerated from thetissue cultures are also part of the invention.

Also included in this invention are methods for producing a corn plantproduced by crossing the inbred line MN5 with itself or another cornline. When crossed with itself, i.e. crossed with another inbred lineMN5 plant or self pollinated, the inbred line MN5 will be conserved.When crossed with another, different corn line, an F₁ hybrid seed isproduced. F₁ hybrid seeds and plants produced by growing said hybridseeds are included in the present invention. A method for producing a F₁hybrid corn seed comprising crossing inbred line MN5 corn plant with adifferent corn plant and harvesting the resultant hybrid corn seed arealso part of the invention. The hybrid corn seed produced by the methodcomprising crossing inbred line MN5 corn plant with a different cornplant and harvesting the resultant hybrid corn seed are included in theinvention, as are included the hybrid corn plant or parts thereof, seedsincluded, produced by growing said hybrid corn seed.

In another embodiment, this invention relates to a method for producingthe inbred line MN5 from a collection of seeds, the collectioncontaining both inbred line MN5 seeds and hybrid seeds having MN5 as aparental line. Such a collection of seed might be a commercial bag ofseeds. Said method comprises planting the collection of seeds. Whenplanted, the collection of seeds will produce inbred line MN5 plantsfrom inbred line MN5 seeds and hybrid plant from hybrid seeds. Theplants having all the physiological and morphological characteristics ofcorn inbred line MN5 or having a decreased vigor compared to the otherplants grown from the collection of seeds are identified as inbred lineMN5 parent plants. Said decreased vigor is due to the inbreedingdepression effect and can be identified for example by a less vigorousappearance for vegetative and/or reproductive characteristics includingshorter plant height, small ear size, ear and kernel shape, ear color orother characteristics. As previously mentioned, if the inbred line MN5is self-pollinated, the inbred line MN5 will be preserved, therefore,the next step is controlling pollination of the inbred parent plants ina manner which preserves the homozygosity of said inbred line MN5 parentplant, and the final step is to harvest the resultant seed.

This invention also relates to methods for producing other inbred cornlines derived from inbred corn line MN5 and to the inbred corn linesderived by the use of those methods.

In another aspect, the present invention provides transformed MN5 inbredcorn line or parts thereof that have been transformed so that itsgenetic material contains one or more transgenes, preferably operablylinked to one or more regulatory elements. Also, the invention providesmethods for producing a corn plant containing in its genetic materialone or more transgenes, preferably operably linked to one or moreregulatory elements, by crossing transformed MN5 inbred corn line witheither a second plant of another corn line, or a non-transformed cornplant of the inbred line MN5, so that the genetic material of theprogeny that results from the cross contains the transgene(s),preferably operably linked to one or more regulatory elements. Theinvention also provides methods for producing a corn plant that containsin its genetic material one or more transgene(s), wherein the methodcomprises crossing the inbred corn line MN5 with a second plant ofanother corn line which contains one or more transgene(s) operablylinked to one or more regulatory element(s) so that the genetic materialof the progeny that results from the cross contains the transgene(s)operably linked to one or more regulatory element(s). Transgenic cornplants, or parts thereof produced by the method are in the scope of thepresent invention.

More specifically, the invention comprises methods for producing malesterile corn plants, male fertile corn plants, herbicide resistant cornplants, insect resistant corn plants, disease resistant corn plants,water stress tolerant corn plants, or plants with modified, inparticular decreased, phytate content, plants with modified waxy and/oramylose starch content, plants with modified protein content, plantswith modified oil content or profile, plants with increaseddigestibility or plants with increased nutritional quality. Said methodscomprise transforming the inbred line MN5 corn plant with nucleic acidmolecules that confer male sterility, male fertility, herbicideresistance, insect resistance, disease resistance, water stresstolerance, or that can modify the phytate, the waxy and/or amylosestarches, the protein or the oil contents, the digestibility or thenutritional qualities, respectively. The transformed corn plantsobtained from the provided methods, including male sterile corn plants,male fertile corn plants, herbicide resistant corn plants, insectresistant corn plants, disease resistant corn plants, water stresstolerant corn plants, plants with modified phytate, waxy and/or amylosestarches, protein or oil contents, plants with increased digestibilityand plants with increased nutritional quality are included in thepresent invention. For the present invention and the skilled artisan,disease is understood to be fungal disease, viral disease, bacterialdisease or other plant pathogenic diseases and disease resistant plantwill encompass plants resistant to fungal, viral, bacterial and otherplant pathogens.

Also included in the invention are methods for producing a corn plantcontaining in its genetic material one or more transgenes involved withfatty acid metabolism, carbohydrate metabolism, and starch content suchas waxy starch or increased amylose starch. The transgenic corn plantsproduced by these methods are also part of the invention.

In another aspect, the present invention provides for methods ofintroducing one or more desired trait(s) into the corn line MN5 andplants obtained from such methods. The desired trait(s) may be, but notexclusively, a single gene, preferably a dominant but also a recessiveallele. Preferably, the transferred gene or genes will confer suchtraits as male sterility, herbicide resistance, insect resistance,resistance for bacterial, fungal, or viral disease, male fertility,water stress tolerance, enhanced nutritional quality, modified waxycontent, modified amylose content, modified protein content, modifiedoil content, enhanced plant quality, enhanced digestibility andindustrial usage. The gene or genes may be naturally occurring maizegene(s) or transgene(s) introduced through genetic engineeringtechniques. The method for introducing the desired trait(s) ispreferably a backcrossing process making use of a series of backcrossesto the inbred corn line MN5 during which the desired trait(s) ismaintained by selection.

When using a transgene, the trait is generally not incorporated intoeach newly developed line such as MN5 by direct transformation. Rather,the more typical method used by breeders of ordinary skill in the art toincorporate the transgene is to take a line already carrying thetransgene and to use such line as a donor line to transfer the transgeneinto the newly developed line. The same would apply for a naturallyoccurring trait. The backcross breeding process comprises the followingsteps: (a) crossing the inbred line MN5 plants with plants of anotherline that comprise the desired trait(s), (b) selecting the F₁ progenyplants that have the desired trait(s); (c) crossing the selected F₁progeny plants with the inbred line MN5 plants to produce backcrossprogeny plants; (d) selecting for backcross progeny plants that have thedesired trait(s) and physiological and morphological characteristics ofcorn inbred line MN5 to produce selected backcross progeny plants; and(e) repeating steps (c) and (d) one, two, three, four, five six, seven,eight, nine or more times in succession to produce selected, second,third, fourth, fifth, sixth, seventh, eighth, ninth or higher backcrossprogeny plants that comprise the desired trait(s) and all thephysiological and morphological characteristics of corn inbred line MN5as listed in Table 1. The corn plants produced by the methods are alsopart of the invention. Backcrossing breeding methods, well known to oneskilled in the art of plant breeding will be further developed insubsequent parts of the specification.

In another aspect of the invention, corn inbred MN5 may be used as aparent, or a single gene conversion or a transgenic inbred line of MN5,such as MN5 BtMon810, may be used as a parent and may be crossed withanother corn line including but not limited to the following corn lines:CB1, KW4773, CC2, BB14 and RBO1. Corn inbred MN5 may be used as a parentor a single gene conversion or a transgenic inbred line of MN5 such asMN5 BtMon810 may be used as a parent and may be crossed with anothercorn line including but not limited to the following corn lines: singlegene conversions or transgenic inbred lines of CB1, such as CB1 RR,single gene conversions or transgenic inbred lines of KW4773, such asKW4773RR, single gene conversions or transgenic inbred lines of CC2,such as CC2RR, single gene conversions or transgenic inbred lines ofKW4U110, such as KW4U110RR, and single gene conversions or transgenicinbred lines of LH311, such as LH311RR. Preferably, the single geneconversions or transgenic inbred lines will confer such traits as malesterility, herbicide resistance, insect resistance, resistance forbacterial, fungal, or viral disease, male fertility, water stresstolerance, enhanced nutritional quality, modified waxy content, modifiedamylose content, modified protein content, modified oil content,enhanced plant quality, enhanced digestibility and industrial usage. Thegene or genes may be naturally occurring maize gene(s) or transgene(s)introduced through genetic engineering techniques. The hybrid cornplants having corn inbred MN5 or a single gene conversion or atransgenic inbred line of MN5 as a parental line and having another,different, corn line as a second parental line as discussed above arecomprised in the present invention.

In a preferred embodiment, the present invention provides methods forincreasing and producing inbred line MN5 seed, whether by crossing afirst inbred parent corn plant with a second inbred parent corn plantand harvesting the resultant corn seed, wherein both said first andsecond inbred corn plant are the inbred line MN5 or by planting aninbred corn seed of the inbred corn line MN5, growing an inbred line MN5plant from said seed, controlling a self pollination of the plant wherethe pollen produced by the grown inbred line MN5 plant pollinates theovules produced by the very same inbred line MN5 grown plant andharvesting the resultant seed.

The invention further provides methods for developing corn plants in acorn plant breeding program using plant breeding techniques includingrecurrent selection, backcrossing, pedigree breeding, molecular marker(Isozyme Electrophoresis, Restriction Fragment Length Polymorphisms(RFLPs), Randomly Amplified Polymorphic DNAs (RAPDs), Arbitrarily PrimedPolymerase Chain Reaction (AP-PCR), DNA Amplification Fingerprinting(DAF), Sequence Characterized Amplified Regions (SCARs). AmplifiedFragment Length Polymorphisms (AFLPs), and Simple Sequence Repeats(SSRs) which are also referred to as Microsatellites, etc.) enhancedselection, genetic marker enhanced selection and transformation. Cornseeds, plants, and parts thereof produced by such breeding methods arealso part of the invention.

In addition to the exemplary aspects and embodiments described above,further aspects and embodiments will become apparent by study of thefollowing descriptions.

DEFINITIONS

In the description and tables which follow, a number of terms are used.In order to provide a clear and consistent understanding of thespecification and claims, including the scope to be given such terms,the following definitions are provided:

Allele. The allele is any of one or more alternative forms of a gene,all of which relate to one trait or characteristic. In a diploid cell ororganism, the two alleles of a given gene occupy corresponding loci on apair of homologous chromosomes.

Backcrossing. Backcrossing is a process in which a breeder repeatedlycrosses hybrid progeny back to one of the parents, for example, a firstgeneration hybrid F₁ with one of the parental genotype of the F₁ hybrid.

Collection of seeds. In the context of the present invention acollection of seeds will be a grouping of seeds mainly containingsimilar kind of seeds, for example hybrid seeds having the inbred lineof the invention as a parental line, but that may also contain, mixedtogether with this first kind of seeds, a second, different kind ofseeds, of one of the inbred parent lines, for example the inbred line ofthe present invention. A commercial bag of hybrid seeds having theinbred line of the invention as a parental line and containing also theinbred line seeds of the invention would be, for example such acollection of seeds.

Daily heat unit value. The daily heat unit value is calculated asfollows: (the maximum daily temperature+the minimum daily temperature)/2minus 50. All temperatures are in degrees Fahrenheit. The maximumtemperature threshold is 86 degrees, if temperatures exceed this, 86 isused. The minimum temperature threshold is 50 degrees, if temperaturesgo below this, 50 is used.

Decreased vigor. A plant having a decreased vigor in the presentinvention is a plant that, compared to other plants has a less vigorousappearance for vegetative and/or reproductive characteristics includingshorter plant height, small ear size, ear and kernel shape, ear color orother characteristics.

Dropped Ears. This is a measure of the number of dropped ears per plot,and represents the percentage of plants that dropped an ear prior toharvest.

Dry down. This is the rate at which a hybrid will reach acceptableharvest moisture

Ear Height. The ear height is a measure from the ground to the ear nodeattachment, and is measured in centimeters.

Essentially all of the physiological and morphological characteristics.A plant having essentially all of the physiological and morphologicalcharacteristics means a plant having the physiological and morphologicalcharacteristics of the recurrent parent, except for the characteristicsderived from the converted gene.

GDU pollen. The number of heat units from planting until 50% of theplants in the hybrid are shedding pollen.

GDU Silk. The GDU silk (=heat unit silk) is the number of growing degreeunits (GDU) or heat units required for an inbred line or hybrid to reachsilk emergence from the time of planting. Growing degree units arecalculated by the Barger Method, where the heat units for a 24-hourperiod are: GDU=((Max Temp+Min Temp)/2)−50 The highest maximum used is86° F. and the lowest minimum used is 50° F. For each hybrid, it takes acertain number of GDUs to reach various stages of plant development.GDUs are a way of measuring plant maturity. GDUs can also relate tostages of growth for an inbred line.

Harvest Aspect. This is a visual rating given the day of harvest or theprevious day. Hybrids are rated 1 (poorest) to 9 (best) with poorerscores given for poor plant health, visible signs of fungal infection,poor plant intactness characterized by missing leaves, tassels, or othervegetative parts, or a combination of these traits.

Herbicide resistant or tolerant. A plant containing anyherbicide-resistant gene or any DNA molecule or construct (or replicatethereof) which is not naturally occurring in the plant which results inincrease tolerance to any herbicide including, imidazoline,sulfonylurea, glyphosate, glufosinate, L-phosphinothricin, triazine andbenzonitrile. For purposes of this definition, a DNA molecule orconstruct shall be considered to be naturally occurring if it exists ina plant at a high enough frequency to provide herbicide resistancewithout further selection and/or if it has not bee produced as a resultof tissue culture selection, mutagenesis, genetic engineering usingrecombinant DNA techniques or other in vitro or in vivo modification tothe plant.

HTU. HTU is the summation of the daily heat unit value calculated fromplanting to harvest.

Inbreeding depression. The inbreeding depression is the loss ofperformance of the inbreds due to the effect of inbreeding, i.e. due tothe mating of relatives or to self-pollination. It increases thehomozygous recessive alleles leading to plants which are weaker andsmaller and having other less desirable traits.

Late plant greenness. Similar to a stay green rating. This is a visualassessment given at around the dent stage but typically a few weeksbefore harvest to characterize the degree of greenness left in theleaves. Plants are rated from 1 (poorest) to 9 (best) with poorer scoresgiven for plants that have more non-green leaf tissue typically due toearly senescence or from disease.

MN RM. This represents the Minnesota Relative Maturity Rating (MN RM)for the hybrid and is based on the harvest moisture of the grainrelative to a standard set of checks of previously determined MN RMrating. Regression analysis is used to compute this rating.

Moisture. The moisture is the actual percentage moisture of the grain atharvest.

Plant Cell. Plant cell, as used herein includes plant cells whetherisolated, in tissue culture, or incorporated in a plant or plant part.

Plant habit. This is a visual assessment assigned during the latevegetative to early reproductive stages to characterize the plants leafhabit. It ranges from decumbent with leaves growing horizontally fromthe stalk to a very upright leaf habit, with leaves growing nearvertically from the stalk.

Plant Height. This is a measure of the height of the hybrid from theground to the tip of the tassel, and is measured in centimeters.

Plant intactness. This is a visual assessment assigned to a hybrid orinbred at or close to harvest to indicate the degree that the plant hassuffered disintegration through the growing season. Plants are ratedfrom 1 (poorest) to 9 (best) with poorer scores given for plants thathave more of their leaf blades missing.

Plant Part. As used herein, the term “plant part” includes leaves,stems, roots, seeds, grains, embryos, pollens, ovules, flowers, ears,cobs, husks, stalks, root tips, anthers, silk, tissue, cells and thelike.

Pollen shed. This is a visual rating assigned at flowering to describethe abundance of pollen produced by the anthers. Inbreds are rated 1(poorest) to 9 (best) with the best scores for inbreds with tassels thatshed more pollen during anthesis.

Post-anthesis Root Lodging. This is a percentage plants that root lodgeafter anthesis: plants that lean from the vertical axis at anapproximately 30° angle or greater.

Pre-anthesis Brittle Snapping. This is a percentage of “snapped” plantsfollowing severe winds prior to anthesis

Pre-anthesis Root Lodging. This is a percentage plants that root lodgeprior to anthesis: plants that lean from the vertical axis at anapproximately 30° angle or greater.

Predicted RM. This trait for a hybrid, predicted relative maturity (RM),is based on the harvest moisture of the grain. The relative maturityrating is based on a known set of checks and utilizes conventionalmaturity such as the Comparative Relative Maturity Rating System or itssimilar, the Minnesota Relative Maturity Rating System.

Quantitative Trait Loci (QTL) Quantitative trait loci refer to geneticloci that control to some degree numerically representable traits thatare usually continuously distributed.

Regeneration. Regeneration refers to the development of a plant fromtissue culture.

Root Lodging. The root lodging is the percentage of plants that rootlodge; i.e., those that lean from the vertical axis at an approximate30° angle or greater would be counted as root lodged.

Seed quality. This is a visual rating assigned to the kernels of theinbred. Kernels are rated 1 (poorest) to 9 (best) with poorer scoresgiven for kernels that are very soft and shriveled with splitting of thepericarp visible and better scores for fully formed kernels.

Seedling Vigor. This is the vegetative growth after emergence at theseedling stage, approximately five leaves.

Silking ability. This is a visual assessment given during flowering.Plants are rated on the amount and timing of silk production. Plants arerated from 1 (poorest) to 9 (best) with poorer scores given for plantsthat produce very little silks that are delayed past pollen shed.

Single gene converted. Single gene converted or conversion plants refersto plants which are developed by a plant breeding technique calledbackcrossing wherein essentially all the morphological and physiologicalcharacteristics of an inbred are recovered in addition to the singlegene transferred into the inbred via the backcrossing technique or viagenetic engineering. This also includes multiple transference of singlegenes.

Stalk Lodging. This is the percentage of plants that stalk lodge, i.e.,stalk breakage, as measured by either natural lodging or pushing thestalks and determining the percentage of plants that break off below theear. This is a relative rating of a hybrid to other hybrids forstandability.

Standability. A term referring to the how well a plant remains uprighttowards the end of the growing season. Plants with excessive stalkbreakage and/or root lodging would be considered to have poorstandability.

Stay Green. Stay green is the measure of plant health near the time ofblack layer formation (physiological maturity). A high score indicatesbetter late-season plant health.

Transgenic. Where an inbred line has been converted to contain one ormore transgenes by single gene conversion or by direct transformation.

Variety. A plant variety as used by one skilled in the art of plantbreeding means a plant grouping within a single botanical taxon of thelowest known rank which can be defined by the expression of thecharacteristics resulting from a given genotype or combination ofphenotypes, distinguished from any other plant grouping by theexpression of at least one of the said characteristics and considered asa unit with regard to its suitability for being propagated unchanged(International convention for the protection of new varieties of plants)

Yield (Bushels/Acre). The yield is the actual yield of the grain atharvest adjusted to 15.5% moisture.

DETAILED DESCRIPTION OF THE INVENTION

Inbred corn line MN5 is a yellow dent corn inbred with superiorcharacteristics, and provides an excellent male parental line in crossesfor producing first generation (F₁) hybrid corn. Inbred corn line MN5 isbest adapted to the East, Central and Western regions of the UnitedStates Corn Belt commonly referred to as Zones 6, 7 and 8. MN5 can beused to produce hybrids having a relative maturity of approximately 108to 115 days on the Comparative Relative Maturity Rating System forharvest moisture of grain. Inbred corn line MN5 shows very good seedlingvigor. MN5 sheds an abundant volume of pollen as a male parent in theproduction of F₁ hybrids. MN5 contributes excellent health to the F₁hybrids which results in strong stalks and good intactness in the F₁hybrids along with a very uniform and an appealing look at harvest. Thisin turn allows farmers to harvest more efficiently and reduces fieldlosses compared to hybrids that have inferior stalks and poor planthealth. MN5 also contributes an excellent yield to moisture ratio to theF₁ hybrids when compared to other Mo17/Lancaster based inbreds. The highyield to moisture ratio and level of production directly correlate to ahigher income per acre for the farmer. MN5 provides the F1 hybrid withhigh levels of disease tolerance and excellent drought and stresstolerance as observed in the testing locations where these conditionswere present. MN5 contributes very good harvest appearance to the F₁hybrids which relates directly to higher levels of plant intactness forthe farmer at harvest. This plant intactness improves harvestefficiency.

MN5 is similar to Mo17, however, there are numerous differencesincluding the fact that MN5 contributes substantially improved planthealth and stalk characteristics to the F₁ hybrid than Mo17. MN5contributes to appreciably lower levels of stalk lodging to hybrids thanother commonly used Lancaster or Mo17 derived inbreds such as LH287.Hybrids with MN5 have significantly improved intactness ratings whichcorrelate to improved harvestability. MN5 has a plant height of 230 cmwith an average ear insertion of 75 cm. The MN5 consistency of earheight is transferred to the F₁ hybrids in the majority of crosses andis exhibited in an eye appealing appearance, especially at harvest time.MN5 is a medium-full season inbred and is very well adapted for use as amale in seed production. The tassel sheds a high volume of pollen. Heatunits to 50% pollen shed are approximately 1470 and to 50% silk areapproximately 1393 as measured near Champaign, Illinois in 2005.

MN5 is an inbred line with very high yield potential in hybrids. Hybridswith MN5 as one parental line produce very uniform, large ears with ahigh kernel row number. Often these hybrid combinations result in plantswhich are appreciably better than average for overall health, rootstrength, stalk strength and plant intactness when compared to inbredlines of similar maturity and family background.

Some of the criteria used to select ears in various generations include:yield, stalk quality, root quality, disease tolerance with emphasis ongrey leaf spot, test weight, late season plant greenness, late seasonplant intactness, ear retention, ear height, pollen shedding ability,silking ability, and corn borer tolerance. The emergence vigor inhybrids with MN5 tends to consistently demonstrate above average vigor.During the development of the line, crosses were made to inbred testersfor the purpose of estimating the line's general and specific combiningability, and evaluations were run by the Champaign, Ill. ResearchStation. The inbred was evaluated further as a line and in numerouscrosses by the Champaign station and other research stations across theCorn Belt. The inbred has proven to have a good combining ability inhybrid combinations.

The inbred line has shown uniformity and stability for the traits,within the limits of environmental influence for the traits. MN5 wasdeveloped through the dihaploid breeding method. It has beenself-pollinated a sufficient number of generations with carefulattention to uniformity of plant type. The line has been increased withcontinued observation for uniformity. No variant traits have beenobserved or are expected in MN5.

Inbred corn line MN5 has the following morphologic and othercharacteristics (based primarily on data collected at Champaign, Ill.).

TABLE 1 VARIETY DESCRIPTION INFORMATION Type: MN5 is a yellow, dent corninbred Region where developed: Champaign, Illinois Maturity: Days HeatUnits From emergence to 50% of plants in silk: 75 1393 From emergence to50% of plants in pollen: 76 1417 Heat Units: = GDU = ((Max Temp + MinTemp)/2) − 50 Plant: Plant Height to tassel tip: 230 cm Ear Height tobase of top ear: 75 cm Average Length of Top Ear Internode: 20 cmAverage number of Tillers: 0 Average Number of Ears per Stalk: 1.3Anthocyanin of Brace Roots: Absent Leaf: Width of Ear Node Leaf: 9.5 cmLength of Ear Node Leaf: 77 cm Number of leaves above top ear: 4 LeafAngle from 2nd Leaf above ear at 30° anthesis to Stalk above leaf: LeafColor: Munsell 5GY-4/4 Leaf Sheath Pubescence (Rated on scale 4 from 1 =none to 9 = like peach fuzz): Marginal Waves (Rated on scale from 5 1 =none to 9 = many): Longitudinal Creases (Rated on scale from 3 1 = noneto 9 = many): Tassel: Number of Lateral Branches: 3 Branch Angle fromCentral Spike: 30 Degrees Tassel Length (from top leaf collar 54 cm totassel top): Pollen Shed (RateD on scale from 0 = male 6 sterile to 9 =heavy shed): Anther Color: Munsell 10Y-8/6 Glume Color: Munsell 5 GY 6/8Tassel Glume Bands Color: Absent Ear: (Unhusked Data) Silk Color (3 daysafter emergence): Munsell 10RP 4/8 Fresh Husk Color (25 days afterMunsell 5GY 7/8 50% silking): Dry Husk Color (65 days after 50%silking): Munsell 5Y 8/10 Position of Ear: Upright Husk Tightness (Ratedon scale from 3 1 = very loose to 9 = very tight): Husk Extension atharvest: Moderate Husk Extension-Ear Tip Covered Ear: (Husked Ear Data)Ear Length: 13.5 cm Ear Diameter at mid-point: 41.3 mm Ear Weight: 108.2gm Number of Kernel Rows: 14–16 Row Alignment: Straight Shank Length:14.9 cm Ear Taper: Cylindrical Kernel: (Dried) Kernel Length: 10.6 mmKernel Width: 8.3 mm Kernel Thickness: 5.6 mm Round Kernels (ShapeGrade): 68.5% Aleurone Color: White Hard Endosperm Color: Munsell 10YR7/12 Endosperm Type: Dent Weight per 100 kernels (unsized sample): 28 gmCob: Cob Diameter at Mid-Point: 24 mm Cob Color: Red Agronomic Traits:Stay Green (at 65 days after anthesis) (Rated 7 on scale from 1 = worstto 9 = excellent): Dropped Ears (at 65 days after anthesis): 0%Pre-anthesis Brittle Snapping: 0% Pre-anthesis Root Lodging: 0%Post-anthesis Root Lodging 0% (at 65 days after anthesis): Yield ofInbred Per Se 46 Bu/Acre (at 12–13% grain moisture):

Further Embodiments of the Invention

This invention is also directed to methods for producing a corn plant bycrossing a first parent corn plant with a second parent corn plantwherein either the first or second parent corn plant is an inbred cornplant of the line MN5. Further, both first and second parent corn plantscan come from the inbred corn line MN5. When self-pollinated, or crossedwith another inbred line MN5 plant, the inbred line MN5 will be stablewhile when crossed with another, different corn line, an F₁ hybrid seedis produced.

An inbred line has been produced through several cycles ofself-pollination and is therefore to be considered as a homozygous line.An inbred line can also be produced though the dihaploid system whichinvolves doubling the chromosomes from a haploid plant thus resulting inan inbred line that is genetically stable (homozygous) and can bereproduced without altering the inbred line. A hybrid variety isclassically created through the fertilization of an ovule from an inbredparental line by the pollen of another, different inbred parental line.Due to the homozygous state of the inbred line, the produced gametescarry a copy of each parental chromosome. As both the ovule and thepollen bring a copy of the arrangement and organization of the genespresent in the parental lines, the genome of each parental line ispresent in the resulting F₁ hybrid, theoretically in the arrangement andorganization created by the plant breeder in the original parental line.

As long as the homozygosity of the parental lines is maintained, theresulting hybrid cross is stable. The F₁ hybrid is then a combination ofphenotypic characteristics issued from two arrangement and organizationof genes, both created by a man skilled in the art through the breedingprocess.

Still further, this invention also is directed to methods for producingan inbred corn line MN5-derived corn plant by crossing inbred corn lineMN5 with a second corn plant and growing the progeny seed, and repeatingthe crossing and growing steps with the inbred corn line MN5-derivedplant from 0 to 7 times. Thus, any such methods using the inbred cornline MN5 are part of this invention: selfing, backcrosses, hybridproduction, crosses to populations, and the like. All plants producedusing inbred corn line MN5 as a parent are within the scope of thisinvention, including plants derived from inbred corn line MN5.Advantageously, the inbred corn line is used in crosses with other,different, corn inbreds to produce first generation (F₁) corn hybridseeds and plants with superior characteristics.

It should be understood that the inbred can, through routinemanipulation of cytoplasmic or other factors, be produced in amale-sterile form. Such embodiments are also contemplated within thescope of the present claims. As used herein, the term plant includesplant cells, plant protoplasts, plant cell tissue cultures from whichcorn plants can be regenerated, plant calli, plant clumps and plantcells that are intact in plants or parts of plants, such as embryos,pollen, ovules, flowers, kernels, seeds, ears, cobs, leaves, husks,stalks, roots, root tips, anthers, silk and the like.

Duncan, et al. (Planta, 1985,165:322-332) indicates that 97% of theplants cultured that produced callus were capable of plant regeneration.Subsequent experiments with both inbreds and hybrids produced 91%regenerable callus that produced plants. In a further study in 1988,Songstad, et al. (Plant Cell Reports, 7:262-265) reports several mediaadditions that enhance regenerability of callus of two inbred lines.Other published reports also indicated that “nontraditional” tissues arecapable of producing somatic embryogenesis and plant regeneration. K. V.Rao et al., (Maize Genetics Cooperation Newsletter, 1986, 60:64-65)refer to somatic embryogenesis from glume callus cultures and B. V.Conger, et al. (Plant Cell Reports, 1987, 6:345-347) indicate somaticembryogenesis from the tissue cultures of corn leaf segments. Thus, itis clear from the literature that the state of the art is such thatthese methods of obtaining plants are routinely used and have a veryhigh rate of success.

Tissue culture of corn is also described in European Patent Application,publication 160, 390 and in Green and Rhodes, Maize for BiologicalResearch, Plant Molecular Biology Association, Charlottesville, Va.,1982, 367-372. Thus, another aspect of this invention is to providecells which upon growth and differentiation produce corn plants havingthe physiological and morphological characteristics of inbred corn lineMN5.

The utility of inbred corn line MN5 also extends to crosses with otherspecies. Commonly, suitable species will be of the family Graminaceae,and especially of the genera Zea, Tripsacum, Coix, Schlerachne,Polytoca, Chionachne, and Trilobachne, of the tribe Maydeae. Potentiallysuitable for crosses with MN5 may be the various varieties of grainsorghum, Sorghum bicolor (L.) Moench.

With the advent of molecular biological techniques that have allowed theisolation and characterization of genes that encode specific proteinproducts, scientists in the field of plant biology developed a stronginterest in engineering the genome of plants to contain and expressforeign genes, or additional, or modified versions of native, orendogenous, genes (perhaps driven by different promoters) in order toalter the traits of a plant in a specific manner. Such foreignadditional and/or modified genes are referred to herein collectively as“transgenes”. Over the last fifteen to twenty years several methods forproducing transgenic plants have been developed, and the presentinvention, in particular embodiments, also relates to transformedversions of the claimed inbred line.

Plant transformation involves the construction of an expression vectorwhich will function in plant cells. Such a vector comprises DNAcomprising a gene under control of, or operatively linked to, aregulatory element (for example, a promoter). The expression vector maycontain one or more such operably linked gene/regulatory elementcombinations. The vector(s) may be in the form of a plasmid, and can beused alone or in combination with other plasmids, to provide transformedcorn plants, using transformation methods as described below toincorporate transgenes into the genetic material of the corn plant(s).

Expression Vectors for Corn Transformation: Marker Genes

Expression vectors include at least one genetic marker, operably linkedto a regulatory element (a promoter, for example) that allowstransformed cells containing the marker to be either recovered bynegative selection, i.e., inhibiting growth of cells that do not containthe selectable marker gene, or by positive selection, i.e., screeningfor the product encoded by the genetic marker. Many commonly usedselectable marker genes for plant transformation are well known in thetransformation arts, and include, for example, genes that code forenzymes that metabolically detoxify a selective chemical agent which maybe an antibiotic or a herbicide, or genes that encode an altered targetwhich is insensitive to the inhibitor. A few positive selection methodsare also known in the art.

One commonly used selectable marker gene for plant transformation is theneomycin phosphotransferase II (nptII) gene, isolated from transposonTn5, which, when placed under the control of plant regulatory signals,confers resistance to kanamycin (Fraley et al., Proc. Natl. Acad. Sci.U.S.A., 80:4803 (1983)). Another commonly used selectable marker gene isthe hygromycin phosphotransferase gene which confers resistance to theantibiotic hygromycin (Vanden Elzen et al., Plant Mol. Biol., 5:299(1985)).

Additional selectable marker genes of bacterial origin that conferresistance to antibiotics include gentamycin acetyl transferase,streptomycin phosphotransferase, and aminoglycoside-3′-adenyltransferase, the bleomycin resistance determinant (Hayford et al., PlantPhysiol. 86:1216 (1988), Jones et al., Mol. Gen. Genet., 210:86 (1987),Svab et al., Plant Mol. Biol. 14:197 (1990), and Hille et al., PlantMol. Biol. 7:171 (1986)). Other selectable marker genes conferresistance to herbicides such as glyphosate, glufosinate or bromoxynil(Comai et al., Nature 317:741-744 (1985), Gordon-Kamm et al., Plant Cell2:603-618 (1990) and Stalker et al., Science 242:419-423 (1988)).

Selectable marker genes for plant transformation that are not ofbacterial origin include, for example, mouse dihydrofolate reductase,plant 5-enolpyruvylshikimate-3-phosphate synthase and plant acetolactatesynthase (Eichholtz et al., Somatic Cell Mol. Genet. 13:67 (1987), Shahet al., Science 233:478 (1986), and Charest et al., Plant Cell Rep.8:643 (1990)).

Another class of marker genes for plant transformation requiresscreening of presumptively transformed plant cells rather than directgenetic selection of transformed cells for resistance to a toxicsubstance such as an antibiotic. These genes are particularly useful toquantify or visualize the spatial pattern of expression of a gene inspecific tissues and are frequently referred to as reporter genesbecause they can be fused to a gene or gene regulatory sequence for theinvestigation of gene expression. Commonly used genes for screeningpresumptively transformed cells include beta-glucuronidase (GUS),beta-galactosidase, luciferase, and chloramphenicol acetyltransferase(Jefferson, R. A., Plant Mol. Biol. Rep. 5:387 (1987), Teeri et al.,EMBO J. 8:343 (1989), Koncz et al., Proc. Natl. Acad. Sci U.S.A. 84:131(1987), and DeBlock et al., EMBO J. 3:1681 (1984). Another approach tothe identification of relatively rare transformation events has been useof a gene that encodes a dominant constitutive regulator of the Zea maysanthocyanin pigmentation pathway (Ludwig et al., Science 247:449(1990)).

In vivo methods for visualizing GUS activity that do not requiredestruction of plant tissue are also available. However, these in vivomethods for visualizing GUS activity have not proven useful for recoveryof transformed cells because of low sensitivity, high fluorescentbackgrounds and limitations associated with the use of luciferase genesas selectable markers.

A gene encoding Green Fluorescent Protein (GFP) has been utilized as amarker for gene expression in prokaryotic and eukaryotic cells (Chalfieet al., Science 263:802 (1994)). GFP and mutants of GFP may be used asscreenable markers.

Expression Vectors for Corn Transformation: Promoters

Genes included in expression vectors must be driven by nucleotidesequence comprising a regulatory element, for example, a promoter.Several types of promoters are now well known in the transformationarts, as are other regulatory elements that can be used alone or incombination with promoters.

As used herein, “promoter” includes reference to a region of DNAupstream from the start of transcription and involved in recognition andbinding of RNA polymerase and other proteins to initiate transcription.A “plant promoter” is a promoter capable of initiating transcription inplant cells. Examples of promoters under developmental control includepromoters that preferentially initiate transcription in certain organs,such as leaves, roots, seeds and tissues such as fibers, xylem vessels,tracheids, or sclerenchyma. Such promoters are referred to as“tissue-preferred”. Promoters which initiate transcription only incertain tissue are referred to as “tissue-specific”. A “cell type”specific promoter primarily drives expression in certain cell types inone or more organs, for example, vascular cells in roots or leaves. An“inducible” promoter is a promoter which is under environmental control.Examples of environmental conditions that may effect transcription byinducible promoters include anaerobic conditions or the presence oflight. Tissue-specific, tissue-preferred, cell type specific, andinducible promoters constitute the class of “non-constitutive”promoters. A “constitutive” promoter is a promoter which is active undermost environmental conditions.

A. Inducible Promoters—An inducible promoter is operably linked to agene for expression in corn. Optionally, the inducible promoter isoperably linked to a nucleotide sequence encoding a signal sequencewhich is operably linked to a gene for expression in corn. With aninducible promoter the rate of transcription increases in response to aninducing agent. Any inducible promoter can be used in the instantinvention. See Ward et al., Plant Mol. Biol. 22:361-366 (1993).Exemplary inducible promoters include, but are not limited to, that fromthe ACEI system which responds to copper (Mett et al., Proc. Natl. Acad.Sci. U.S.A. 90:4567-4571 (1993)); In2 gene from maize which responds tobenzenesulfonamide herbicide safeners (Gatz et al., Mol. Gen. Genetics243:32-38 (1994)) or Tet repressor from Tn10 (Gatz et al., Mol. Gen.Genetics 227:229-237 (1991)). A particularly preferred induciblepromoter is a promoter that responds to an inducing agent to whichplants do not normally respond. An exemplary inducible promoter is theinducible promoter from a steroid hormone gene, the transcriptionalactivity of which is induced by a glucocorticosteroid hormone (Schena etal., Proc. Natl. Acad. Sci. U.S.A. 88:0421 (1991)).

B. Constitutive Promoters—A constitutive promoter is operably linked toa gene for expression in corn or the constitutive promoter is operablylinked to a nucleotide sequence encoding a signal sequence which isoperably linked to a gene for expression in corn. Many differentconstitutive promoters can be utilized in the instant invention.Exemplary constitutive promoters include, but are not limited to, thepromoters from plant viruses such as the 35S promoter from CaMV (Odellet al., Nature 313:810-812 (1985)) and the promoters from such genes asrice actin (McElroy et al., Plant Cell 2:163-171 (1990)); ubiquitin(Christensen et al., Plant Mol. Biol. 12:619-632 (1989) and Christensenet al., Plant Mol. Biol. 18:675-689 (1992)); pEMU (Last et al., Theor.Appl. Genet. 81:581-588 (1991)); MAS (Velten et al., EMBO J. 3:2723-2730(1984)) and maize H3 histone (Lepetit et al., Mol. Gen. Genetics231:276-285 (1992) and Atanassova et al., Plant Journal 2 (3): 291-300(1992)).

The ALS promoter, Xba1/NcoI fragment 5′ to the Brassica napus ALS3structural gene (or a nucleotide sequence similarity to said Xba1/Ncolfragment), represents a particularly useful constitutive promoter. SeePCT application WO96/30530.

C. Tissue-specific or Tissue-preferred Promoters—A tissue-specificpromoter is operably linked to a gene for expression in corn.Optionally, the tissue-specific promoter is operably linked to anucleotide sequence encoding a signal sequence which is operably linkedto a gene for expression in corn. Plants transformed with a gene ofinterest operably linked to a tissue-specific promoter produce theprotein product of the transgene exclusively, or preferentially, in aspecific tissue.

Any tissue-specific or tissue-preferred promoter can be utilized in theinstant invention. Exemplary tissue-specific or tissue-preferredpromoters include, but are not limited to, a root-preferred promoter,such as that from the phaseolin gene (Murai et al., Science 23:476-482(1983) and Sengupta-Gopalan et al., Proc. Natl. Acad. Sci. U.S.A.82:3320-3324 (1985)); a leaf-specific and light-induced promoter such asthat from cab or rubisco (Simpson et al., EMBO J. 4(11):2723-2729 (1985)and Timko et al., Nature 318:579-582 (1985)); an anther-specificpromoter such as that from LAT52 (Twell et al., Mol. Gen. Genetics217:240-245 (1989)); a pollen-specific promoter such as that from Zm13or a microspore-preferred promoter such as that from apg (Twell et al.,Sex. Plant Reprod. 6:217-224 (1993)).

Signal Sequences for Targeting Proteins to Subcellular Compartments

Transport of protein produced by transgenes to a subcellular compartmentsuch as the chloroplast, vacuole, peroxisome, glyoxysome, cell wall ormitochondrion or for secretion into the apoplast, is accomplished bymeans of operably linking the nucleotide sequence encoding a signalsequence to the 5′ and/or 3′ region of a gene encoding the protein ofinterest. Targeting sequences at the 5′ and/or 3′ end of the structuralgene may determine, during protein synthesis and processing, where theencoded protein is ultimately compartmentalized.

The presence of a signal sequence directs a polypeptide to either anintracellular organelle or subcellular compartment or for secretion tothe apoplast. Many signal sequences are known in the art. See, forexample Becker et al., Plant Mol. Biol. 20:49 (1992), Knox, C., et al.,Plant Mol. Biol. 9:3-17 (1987), Lerner et al., Plant Physiol. 91:124-129(1989), Fontes et al., Plant Cell 3:483-496 (1991), Matsuoka et al.,Proc. Natl. Acad. Sci. 88:834 (1991), Gould et al., J. Cell. Biol.108:1657 (1989), Creissen et al., Plant J. 2:129 (1991), Kalderon, etal., Cell 39:499-509 (1984), Stiefel, et al., Plant Cell 2:785-793(1990).

Foreign Protein Genes and Agronomic Genes

With transgenic plants according to the present invention, a foreignprotein can be produced in commercial quantities. Thus, techniques forthe selection and propagation of transformed plants, which are wellunderstood in the art, yield a plurality of transgenic plants which areharvested in a conventional manner, and a foreign protein then can beextracted from a tissue of interest or from total biomass. Proteinextraction from plant biomass can be accomplished by known methods whichare discussed, for example, by Heney and Orr, Anal. Biochem. 114:92-6(1981).

According to a preferred embodiment, the transgenic plant provided forcommercial production of foreign protein is corn. In another preferredembodiment, the biomass of interest is seed. For the relatively smallnumber of transgenic plants that show higher levels of expression, agenetic map can be generated, primarily via conventional RFLP, PCR andSSR analysis, which identifies the approximate chromosomal location ofthe integrated DNA molecule. For exemplary methodologies in this regard,see Glick and Thompson, Methods in Plant Molecular Biology andBiotechnology CRC Press, Boca Raton 269:284 (1993). Map informationconcerning chromosomal location is useful for proprietary protection ofa subject transgenic plant. If unauthorized propagation is undertakenand crosses made with other germplasm, the map of the integration regioncan be compared to similar maps for suspect plants, to determine if thelatter have a common parentage with the subject plant. Map comparisonswould involve hybridizations, RFLP, PCR, SSR and sequencing, all ofwhich are conventional techniques.

Likewise, by means of the present invention, agronomic genes can beexpressed in transformed plants. More particularly, plants can begenetically engineered to express various phenotypes of agronomicinterest. Exemplary genes implicated in this regard include, but are notlimited to, those categorized below:

1. Genes That Confer Resistance to Pests or Disease and That Encode:

A. Plant disease resistance genes. Plant defenses are often activated byspecific interaction between the product of a disease resistance gene(R) in the plant and the product of a corresponding avirulence (Avr)gene in the pathogen. A plant inbred line can be transformed with clonedresistance gene to engineer plants that are resistant to specificpathogen strains. See, for example Jones et al., Science 266:789 (1994)(cloning of the tomato Cf-9 gene for resistance to Cladosporium fulvum);Martin et al., Science 262:1432 (1993) (tomato Pto gene for resistanceto Pseudomonas syringae pv. tomato encodes a protein kinase); Mindrinoset al., Cell 78:1089 (1994) (Arabidopsis RSP2 gene for resistance toPseudomonas syringae).

B. A Bacillus thuringiensis protein, a derivative thereof or a syntheticpolypeptide modeled thereon. See, for example, Geiser et al., Gene48:109 (1986), who disclose the cloning and nucleotide sequence of a Btalpha-endotoxin gene. Moreover, DNA molecules encoding alpha-endotoxingenes can be purchased from American Type Culture Collection, Manassas,Va., for example, under ATCC Accession Nos. 40098, 67136, 31995 and31998.

C. A lectin. See, for example, the article by Van Damme et al., PlantMolec. Biol. 24:25 (1994), who disclose the nucleotide sequences ofseveral Clivia miniata mannose-binding lectin genes.

D. A vitamin-binding protein such as avidin. See PCT application U.S.Pat. No. 93/06487. The application teaches the use of avidin and avidinhomologues as larvicides against insect pests.

E. An enzyme inhibitor, for example, a protease or proteinase inhibitoror an amylase inhibitor. See, for example, Abe et al., J. Biol. Chem.262:16793 (1987) (nucleotide sequence of rice cysteine proteinaseinhibitor), Huub et al., Plant Molec. Biol. 21:985 (1993) (nucleotidesequence of cDNA encoding tobacco proteinase inhibitor I), Sumitani etal., Biosci. Biotech. Biochem. 57:1243 (1993) (nucleotide sequence ofStreptomyces nitrosporeus alpha-amylase inhibitor).

F. An insect-specific hormone or pheromone such as an ecdysteroid andjuvenile hormone, a variant thereof, a mimetic based thereon, or anantagonist or agonist thereof. See, for example, the disclosure byHammock et al., Nature 344:458 (1990), of baculovirus expression ofcloned juvenile hormone esterase, an inactivator of juvenile hormone.

G. An insect-specific peptide or neuropeptide which, upon expression,disrupts the physiology of the affected pest. For example, see thedisclosures of Pratt et al., Biochem. Biophys. Res. Comm. 163:1243(1989) (an allostatin is identified in Diploptera puntata). See alsoU.S. Pat. No. 5,266,317 to Tomalski et al., who disclose genes encodinginsect-specific, paralytic neurotoxins.

H. An insect-specific venom produced in nature by a snake, a wasp, etc.For example, see Pang et al., Gene 116:165 (1992), for disclosure ofheterologous expression in plants of a gene coding for a scorpioninsectotoxic peptide.

I. An enzyme responsible for a hyper-accumulation of a monoterpene, asesquiterpene, a steroid, a hydroxamic acid, a phenylpropanoidderivative or another non-protein molecule with insecticidal activity.

J. An enzyme involved in the modification, including thepost-translational modification, of a biologically active molecule; forexample, a glycolytic enzyme, a proteolytic enzyme, a lipolytic enzyme,a nuclease, a cyclase, a transaminase, an esterase, a hydrolase, aphosphatase, a kinase, a phosphorylase, a polymerase, an elastase, achitinase and a glucanase, whether natural or synthetic. See PCTapplication WO 93/02197 in the name of Scott et al., which discloses thenucleotide sequence of a callase gene. DNA molecules which containchitinase-encoding sequences can be obtained, for example, from the ATCCunder Accession Nos. 39637 and 67152. See also Kramer et al., InsectBiochem. Molec. Biol. 23:691 (1993), who teach the nucleotide sequenceof a cDNA encoding tobacco hornworm chitinase, and Kawalleck et al.,Plant Molec. Biol. 21:673 (1993), who provide the nucleotide sequence ofthe parsley ubi4-2 polyubiquitin gene.

K. A molecule that stimulates signal transduction. For example, see thedisclosure by Botella et al., Plant Molec. Biol. 24:757 (1994), ofnucleotide sequences for mung bean calmodulin cDNA clones, and Griess etal., Plant Physiol. 104:1467 (1994), who provide the nucleotide sequenceof a maize calmodulin cDNA clone.

L. A hydrophobic moment peptide. See PCT application WO95/16776(disclosure of peptide derivatives of Tachyplesin which inhibit fungalplant pathogens) and PCT application WO95/18855 (teaches syntheticantimicrobial peptides that confer disease resistance).

M. A membrane permease, a channel former or a channel blocker. Forexample, see the disclosure of Jaynes et al., Plant Sci 89:43 (1993), ofheterologous expression of a cecropin-beta, lytic peptide analog torender transgenic tobacco plants resistant to Pseudomonas solanacearum.

N. A viral-invasive protein or a complex toxin derived therefrom. Forexample, the accumulation of viral coat proteins in transformed plantcells imparts resistance to viral infection and/or disease developmenteffected by the virus from which the coat protein gene is derived, aswell as by related viruses. See Beachy et al., Ann. Rev. Phytopathol.28:451 (1990). Coat protein-mediated resistance has been conferred upontransformed plants against alfalfa mosaic virus, cucumber mosaic virus,tobacco streak virus, potato virus X, potato virus Y, tobacco etchvirus, tobacco rattle virus and tobacco mosaic virus. Id.

O. An insect-specific antibody or an immunotoxin derived therefrom.Thus, an antibody targeted to a critical metabolic function in theinsect gut would inactivate an affected enzyme, killing the insect.

P. A virus-specific antibody. See, for example, Tavladoraki et al.,Nature 366:469 (1993), who show that transgenic plants expressingrecombinant antibody genes are protected from virus attack.

Q. A developmental-arrestive protein produced in nature by a pathogen ora parasite. Thus, fungal endo-alpha-1,4-D-polygalacturonases facilitatefungal colonization and plant nutrient release by solubilizing plantcell wall homo-alpha-1,4-D-galacturonase. See Lamb et al., BioTechnology10:1436 (1992). The cloning and characterization of a gene which encodesa bean endopolygalacturonase-inhibiting protein is described by Toubartet al., Plant J. 2:367 (1992).

R. A developmental-arrestive protein produced in nature by a plant. Forexample, Logemann et al., BioTechnology 10:305 (1992), have shown thattransgenic plants expressing the barley ribosome-inactivating gene havean increased resistance to fungal disease.

2. Genes That Confer Resistance to an Herbicide, For Example:

A. An herbicide that inhibits the growing point or meristem, such as animidazolinone or a sulfonylurea. Exemplary genes in this category codefor mutant ALS and AHAS enzyme as described, for example, by Lee et al.,EMBO J. 7:1241 (1988), and Miki et al., Theor. Appl. Genet. 80:449(1990), respectively.

B. Glyphosate (resistance conferred by mutant5-enolpyruvylshikimate-3-phosphate synthase (EPSP) and aroA genes,respectively) and other phosphono compounds such as glufosinate(phosphinothricin acetyl transferase (PAT) and Streptomyceshygroscopicus PAT bar genes), and pyridinoxy or phenoxy propionic acidsand cyclohexones (ACCase inhibitor-encoding genes). See, for example,U.S. Pat. No. 4,940,835 to Shah, et al., which discloses the nucleotidesequence of a form of EPSP which can confer glyphosate resistance. A DNAmolecule encoding a mutant aroA gene can be obtained under ATCCaccession number 39256, and the nucleotide sequence of the mutant geneis disclosed in U.S. Pat. No. 4,769,061 to Comai. European patentapplication No. 0 333 033 to Kumada et al., and U.S. Pat. No. 4,975,374to Goodman et al., disclose nucleotide sequences of glutamine synthetasegenes which confer resistance to herbicides such as L-phosphinothricin.The nucleotide sequence of a PAT gene is provided in Europeanapplication No. 0 242 246 to Leemans et al. DeGreef et al.,BioTechnology 7:61 (1989), describe the production of transgenic plantsthat express chimeric bar genes coding for PAT activity. Exemplary ofgenes conferring resistance to phenoxy propionic acids and cyclohexones,such as sethoxydim and haloxyfop are the Acc1-S1, Acc1-S2 and Acc1-S3genes described by Marshall et al., Theor. Appl. Genet. 83:435 (1992).

C. An herbicide that inhibits photosynthesis, such as a triazine (psbAand gs+ genes) or a benzonitrile (nitrilase gene). Przibilla et al.,Plant Cell 3:169 (1991), describe the transformation of Chlamydomonaswith plasmids encoding mutant psbA genes. Nucleotide sequences fornitrilase genes are disclosed in U.S. Pat. No. 4,810,648 to Stalker, andDNA molecules containing these genes are available under ATCC AccessionNos. 53435, 67441, and 67442. Cloning and expression of DNA coding for aglutathione S-transferase is described by Hayes et al., Biochem. J.285:173 (1992).

3. Genes That Confer or Contribute to a Value-Added Trait, Such as:

A. Modified fatty acid metabolism, for example, by transforming a plantwith an antisense gene of stearyl-ACP desaturase to increase stearicacid content of the plant. See Knutzon et al., Proc. Natl. Acad. Sci.U.S.A. 89:2624 (1992)

B. Increased resistance to high light stress such as photo-oxidativedamages, for example by transforming a plant with a gene coding for aprotein of the Early Light Induced Protein family (ELIP) as described inWO 03074713 in the name of Biogemma.

C. Modified carbohydrate composition effected, for example, bytransforming plants with a gene coding for an enzyme that alters thebranching pattern of starch. See Shiroza et al., J. Bact. 170:810 (1988)(nucleotide sequence of Streptococcus mutants fructosyltransferasegene), Steinmetz et al., Mol. Gen. Genet. 20:220 (1985) (nucleotidesequence of Bacillus subtilis levansucrase gene), Pen et al.,BioTechnology 10:292 (1992) (production of transgenic plants thatexpress Bacillus lichenifonnis α-amylase), Elliot et al., Plant Molec.Biol. 21:515 (1993) (nucleotide sequences of tomato invertase genes),Søgaard et al., J. Biol. Chem. 268:22480 (1993) (site-directedmutagenesis of barley alpha-amylase gene), and Fisher et al., PlantPhysiol. 102:1045 (1993) (maize endosperm starch branching enzyme II).

D. Increased resistance/tolerance to water stress or drought, forexample, by transforming a plant to create a plant having a modifiedcontent in ABA-Water-Stress-Ripening-Induced proteins (ARS proteins) asdescribed in WO 0183753 in the name of Biogemma, or by transforming aplant with a nucleotide sequence coding for a phosphoenolpyruvatecarboxylase as shown in WO02081714. The tolerance of corn to drought canalso be increased by an overexpression of phosphoenolpyruvatecarboxylase (PEPC-C4), obtained, for example from sorghum.

E. Increased content of cysteine and glutathione, useful in theregulation of sulfur compounds and plant resistance against variousstresses such as drought, heat or cold, by transforming a plant with agene coding for an Adenosine 5′ Phosphosulfate as shown in WO 0149855.

F. Increased nutritional quality, for example, by introducing a zeingene which genetic sequence has been modified so that its proteinsequence has an increase in lysine and proline. The increasednutritional quality can also be attained by introducing into the maizeplant an albumin 2S gene from sunflower that has been modified by theaddition of the KDEL peptide sequence to keep and accumulate the albuminprotein in the endoplasmic reticulum.

G. Decreased phytate content: 1) Introduction of a phytase-encoding genewould enhance breakdown of phytate, adding more free phosphate to thetransformed plant. For example, see Van Hartingsveldt et al., Gene127:87 (1993), for a disclosure of the nucleotide sequence of anAspergillus niger phytase gene. 2) A gene could be introduced thatreduced phytate content. In maize, this, for example, could beaccomplished, by cloning and then reintroducing DNA associated with thesingle allele which is responsible for maize mutants characterized bylow levels of phytic acid. See Raboy et al., Maydica 35:383 (1990).

4. Genes that Control Male Sterility

A. Introduction of a deacetylase gene under the control of atapetum-specific promoter and with the application of the chemicalN-Ac-PPT. See international publication WO 01/29237.

B. Introduction of various stamen-specific promoters. See internationalpublications WO 92/13956 and WO 92/13957.

C. Introduction of the barnase and the barstar genes. See Paul et al.,Plant Mol. Biol. 19:611-622, 1992).

Examples of Transgenes

MON810 is the designation given by the Monsanto Company (St. Louis, Mo.)for the transgenic event that, when expressed in maize, produces anendotoxin that is efficacious against the European corn borer, Ostrinianubilalis and certain other Lepidopteran larvae.

MON603, better known as NK603, is the designation for the transgenicevent that, when expressed in maize, allows the use of glyphosate as aweed control agent on the crop. Another transgenic event, GA21, whenexpressed in maize, also allows the use of glyphosate as a weed controlagent on the crop.

MON863 is the designation for the transgenic event that, when expressedin maize, produces an endotoxin that is efficacious against the cornroot worm, Diabrotica vigifera, and certain other Coleopteran larvae.

MON88017 is the transgenic event that, when expressed in maize, allowsthe use of glyphosate as a weed control agent. In addition, this eventproduces an endotoxin that is efficacious against the corn root worm,Diabrotica virgifera, and certain other Coleopteran larvae.

HERCULEX Corn Borer, better known as HX1 or TC1507, is the designationfor the transgenic event that, when expressed in maize, produces anendotoxin that is efficacious against the European corn borer, Ostrinianubilalis, and certain other Lepidopteran larvae. In addition, thetransgenic event was developed to allow the crop to be tolerant to theuse of glufosinate ammonium, the active ingredient in phosphinothricinherbicides.

HERCULEX Root Worm, or DAS59122-7, is the designation for the transgenicevent that, when expressed in maize, produces an endotoxin that isefficacious against the corn root worm, Diabrotica virgifera, andcertain other Coleopteran larvae. In addition, the transgenic event wasdeveloped to allow the crop to be tolerant to the use of glufosinateammonium, the active ingredient in phosphinothricin herbicides.

T25 is the designation for the transgenic event that, when expressed inmaize, allows the crop to be tolerant to the use of glufosinateammonium, the active ingredient in phosphinothricin herbicides.

Methods for Corn Transformation

Numerous methods for plant transformation have been developed, includingbiological and physical plant transformation protocols. See, forexample, Miki et al., “Procedures for Introducing Foreign DNA intoPlants” in Methods in Plant Molecular Biology and Biotechnology, GlickB. R. and Thompson, J. E. Eds. (CRC Press, Inc., Boca Raton, 1993) pages67-88. In addition, expression vectors and in vitro culture methods forplant cell or tissue transformation and regeneration of plants areavailable. See, for example, Gruber et al., “Vectors for PlantTransformation” in Methods in Plant Molecular Biology and Biotechnology,Glick B. R. and Thompson, J. E. Eds. (CRC Press, Inc., Boca Raton, 1993)pages 89-119.

A. Agrobacterium-mediated Transformation—One method for introducing anexpression vector into plants is based on the natural transformationsystem of Agrobacterium. See, for example, Horsch et al., Science227:1229 (1985). A. tumefaciens and A. rhizogenes are plant pathogenicsoil bacteria which genetically transform plant cells. The Ti and Riplasmids of A. tumefaciens and A. rhizogenes, respectively, carry genesresponsible for genetic transformation of the plant. See, for example,Kado, C. I., Crit. Rev. Plant Sci. 10:1 (1991). Descriptions ofAgrobacterium vector systems and methods for Agrobacterium-mediated genetransfer are provided by Gruber et al., supra, Miki et al., supra, andMoloney et al., Plant Cell Reports 8:238 (1989). See also, U.S. Pat. No.5,591,616 issued Jan. 7, 1997.

B. Direct Gene Transfer—Despite the fact the host range forAgrobacterium-mediated transformation is broad, some major cereal cropspecies and gymnosperms have generally been recalcitrant to this mode ofgene transfer, even though some success has recently been achieved inrice and corn. Hiei et al., The Plant Journal 6:271-282 (1994) and U.S.Pat. No. 5,591,616 issued Jan. 7, 1997. Several methods of planttransformation, collectively referred to as direct gene transfer, havebeen developed as an alternative to Agrobacterium-mediatedtransformation.

A generally applicable method of plant transformation ismicroprojectile-mediated transformation wherein DNA is carried on thesurface of microprojectiles measuring 1 to 4 micron. The expressionvector is introduced into plant tissues with a biolistic device thataccelerates the microprojectiles to speeds of 300 to 600 m/s which issufficient to penetrate plant cell walls and membranes. Sanford et al.,Part. Sci. Technol. 5:27 (1987), Sanford, J. C., Trends Biotech. 6:299(1988), Klein et al., BioTechnology 6:559-563 (1988), Sanford, J. C.,Physiol Plant 7:206 (1990), Klein et al., BioTechnology 10:268 (1992).In corn, several target tissues can be bombarded with DNA-coatedmicroprojectiles in order to produce transgenic plants, including, forexample, callus (Type I or Type II), immature embryos, and meristematictissue.

Another method for physical delivery of DNA to plants is sonication oftarget cells. Zhang et al., BioTechnology 9:996 (1991). Alternatively,liposome and spheroplast fusion have been used to introduce expressionvectors into plants. Deshayes et al., EMBO J., 4:2731 (1985), Christouet al., Proc Natl. Acad. Sci. U.S.A. 84:3962 (1987). Direct uptake ofDNA into protoplasts using CaCl₂ precipitation, polyvinyl alcohol orpoly-L-ornithine has also been reported. Hain et al., Mol. Gen. Genet.199:161 (1985) and Draper et al., Plant Cell Physiol. 23:451 (1982).Electroporation of protoplasts and whole cells and tissues have alsobeen described. D'Halluin et al., Plant Cell 4:1495-1505 (1992) andSpencer et al., Plant Mol. Biol. 24:51-61 (1994).

Following transformation of corn target tissues, expression of theabove-described selectable marker genes allows for preferentialselection of transformed cells, tissues and/or plants, usingregeneration and selection methods now well known in the art.

The foregoing methods for transformation would typically be used forproducing a transgenic inbred line. The transgenic inbred line couldthen be crossed, with another (non-transformed or transformed) inbredline, in order to produce a new transgenic inbred line. Alternatively, agenetic trait which has been engineered into a particular corn lineusing the foregoing transformation techniques could be moved intoanother line using traditional backcrossing techniques that are wellknown in the plant breeding arts. For example, a backcrossing approachcould be used to move an engineered trait from a public, non-eliteinbred line into an elite inbred line, or from an inbred line containinga foreign gene in its genome into an inbred line or lines which do notcontain that gene. As used herein, “crossing” can refer to a simple X byY cross, or the process of backcrossing, depending on the context.

When the term inbred corn plant is used in the context of the presentinvention, this also includes any inbred corn plant where one or moredesired trait has been introduced through backcrossing methods, whethersuch trait is a naturally occurring one or a transgenic one.Backcrossing methods can be used with the present invention to improveor introduce one or more characteristic into the inbred. The termbackcrossing as used herein refers to the repeated crossing of a hybridprogeny back to one of the parental corn plants for that inbred. Theparental corn plant which contributes the gene or the genes for thedesired characteristic is termed the nonrecurrent or donor parent. Thisterminology refers to the fact that the nonrecurrent parent is used onetime in the backcross protocol and therefore does not recur. Theparental corn plant to which the gene or genes from the nonrecurrentparent are transferred is known as the recurrent parent as it is usedfor several rounds in the backcrossing protocol (Fehr, 1987).

In a typical backcross protocol, the original inbred of interest(recurrent parent) is crossed to a second inbred (nonrecurrent parent)that carries the gene or genes of interest to be transferred. Theresulting progeny from this cross are then crossed again to therecurrent parent and the process is repeated until a corn plant isobtained wherein all the desired morphological and physiologicalcharacteristics of the recurrent parent are recovered in the convertedplant in addition to the gene or genes transferred from the nonrecurrentparent. It should be noted that some, one, two, three or more,self-pollination and growing of a population might be included betweentwo successive backcrosses. Indeed, an appropriate selection in thepopulation produced by the self-pollination, i.e. selection for thedesired trait and physiological and morphological characteristics of therecurrent parent might be equivalent to one, two or even threeadditional backcrosses in a continuous series without rigorousselection, saving time, money and effort for the breeder. A non limitingexample of such a protocol would be the following: a) the firstgeneration F₁ produced by the cross of the recurrent parent A by thedonor parent B is backcrossed to parent A, b) selection is practiced forthe plants having the desired trait of parent B, c) selected plants areself-pollinated to produce a population of plants where selection ispracticed for the plants having the desired trait of parent B and thephysiological and morphological characteristics of parent A, d) theselected plants are backcrossed one, two, three, four, five or moretimes to parent A to produce selected backcross progeny plantscomprising the desired trait of parent B and the physiological andmorphological characteristics of parent A. Step c) may or may not berepeated and included between the backcrosses of step d.

The selection of a suitable recurrent parent is an important step for asuccessful backcrossing procedure. The goal of a backcross protocol isto alter or substitute one or more trait(s) or characteristic(s) in theoriginal inbred. To accomplish this, a gene or genes of the recurrentinbred is modified or substituted with the desired gene or genes fromthe nonrecurrent parent, while retaining essentially all of the rest ofthe desired genetic, and therefore the desired physiological andmorphological, constitution of the original inbred. The choice of theparticular nonrecurrent parent will depend on the purpose of thebackcross; one of the major purposes is to add some commerciallydesirable, agronomically important trait(s) to the plant. The exactbackcrossing protocol will depend on the characteristic(s) or trait(s)being altered to determine an appropriate testing protocol. Althoughbackcrossing methods are simplified when the characteristic beingtransferred is a single gene and dominant allele, multiple genes andrecessive allele(s) may also be transferred and therefore, backcrossbreeding is by no means restricted to character(s) governed by one or afew genes. In fact the number of genes might be less important that theidentification of the character(s) in the segregating population. Inthis instance it may then be necessary to introduce a test of theprogeny to determine if the desired characteristic(s) has beensuccessfully transferred. Such tests encompass not only visualinspection and simple crossing, but also follow up of thecharacteristic(s) through genetically associated markers and molecularassisted breeding tools. For example, selection of progeny containingthe transferred trait is done by direct selection, visual inspection fora trait associated with a dominant allele, while the selection ofprogeny for a trait that is transferred via a recessive allele, such asthe waxy starch characteristic, require selfing the progeny to determinewhich plant carry the recessive allele(s).

Many single gene traits have been identified that are not regularlyselected for in the development of a new inbred but that can be improvedby backcrossing techniques. Single gene traits may or may not betransgenic, i.e. they may be naturally present in the non recurrentparent, examples of these traits include but are not limited to, malesterility, waxy starch, amylose starch, herbicide resistance, resistancefor bacterial, fungal, or viral disease, insect resistance, malefertility, water stress tolerance, enhanced nutritional quality,industrial usage, increased digestibility yield stability and yieldenhancement. An example of this is the Rp1D gene which controlsresistance to rust fungus by preventing P. sorghi from producing spores.The Rp1D gene is usually preferred over the other Rp genes because it iswidely effective against all races of rust, but the emergence of newraces has lead to the use of other Rp genes comprising, for example, theRp1E, Rp1G, Rp1I, Rp1K or “compound” genes which combine two or more Rpgenes including Rp1GI, Rp1GDJ, etc. These genes are generally inheritedthrough the nucleus. Some known exceptions to this are the genes formale sterility, some of which are inherited cytoplasmically, but stillact as single gene traits. Several of these single gene traits aredescribed in U.S. Pat. Nos. 5,777,196; 5,948,957 and 5,969,212, thedisclosures of which are specifically hereby incorporated by reference.

In 1981, the backcross method of breeding accounted for 17% of the totalbreeding effort for inbred line development in the United States,according to, Hallauer, A. R. et al. (1988) “Corn Breeding” in Corn andCorn Improvement, No. 18, pp. 463-481. The backcross breeding methodprovides a precise way of improving varieties that excel in a largenumber of attributes but are deficient in a few characteristics (Allard,1960, Principles of Plant Breeding, John Wiley & Sons, Inc.). The methodmakes use of a series of backcrosses to the variety to be improvedduring which the character or the characters in which improvement issought is maintained by selection. At the end of the backcrossing, thegene or genes being transferred unlike all other genes will beheterozygous. Selfing after the last backcross produces homozygosity forthis gene pair(s) and, coupled with selection, will result in a varietywith exactly the adaptation, yielding ability and qualitycharacteristics of the recurrent parent but superior to that parent inthe particular characteristic(s) for which the improvement program wasundertaken. Therefore, this method provides the plant breeder with ahigh degree of genetic control of his work.

The method is scientifically exact because the morphological andagricultural features of the improved variety could be described inadvance and because the same variety could, if it were desired, be breda second time by retracing the same steps (Briggs, “Breeding wheatsresistant to bunt by the backcross method”, 1930 Jour. Amer. Soc.Agron., 22: 289-244).

Backcrossing is a powerful mechanism for achieving homozygosity and anypopulation obtained by backcrossing must rapidly converge on thegenotype of the recurrent parent. When backcrossing is made the basis ofa plant breeding program, the genotype of the recurrent parent will bemodified only with regards to genes being transferred, which aremaintained in the population by selection.

Examples of successful backcrosses are the transfer of stem rustresistance from “Hope” wheat to “Bart” wheat and the transfer of buntresistance to “Bart” wheat to create “Bart 38” which has resistance toboth stem rust and bunt. Also highlighted by Allard is the successfultransfer of mildew, leaf spot and wilt resistances in “CaliforniaCommon” alfalfa to create “Caliverde”. This new “Caliverde” varietyproduced through the backcross process is indistinguishable from“California Common” except for its resistance to the three nameddiseases.

One of the advantages of the backcross method is that the breedingprogram can be carried out in almost every environment that will allowthe development of the character being transferred.

The backcross technique is not only desirable when breeding for diseaseresistance but also for the adjustment of morphological characters,color characteristics and simply inherited quantitative characters suchas earliness, plant height and seed size and shape. In this regard, amedium grain type variety, “Calady”, has been produced by Jones andDavis. In dealing with quantitative characteristics, they selected thedonor parent with the view of sacrificing some of the intensity of thecharacter for which it was chosen, i.e. grain size. “Lady Wright”, along grain variety was used as the donor parent and “Coloro”, a shortgrain one as the recurrent parent. After four backcrosses, the mediumgrain type variety “Calady” was produced.

INDUSTRIAL APPLICABILITY

Corn is used as human food, livestock feed, and as raw material inindustry. The food uses of corn, in addition to human consumption ofcorn kernels, include both products of dry- and wet-milling industries.The principal products of corn dry-milling are grits, meal and flour.The corn wet-milling industry can provide corn starch, corn syrups, anddextrose for food use. Corn oil is recovered from corn germ, which is aby-product of both dry- and wet-milling industries.

Corn, including both grain and non-grain portions of the plant, is alsoused extensively as livestock feed, primarily for beef cattle, dairycattle, hogs and poultry. Industrial uses of corn include production ofethanol, corn starch in the wet-milling industry and corn flour in thedry-milling industry. The industrial applications of corn starch andflour are based on functional properties, such as viscosity, filmformation, adhesive properties, and ability to suspend particles. Cornstarch and flour have application in the paper and textile industries.Other industrial uses include applications in adhesives, buildingmaterials, foundry binders, laundry starches, explosives, oil-well mudsand other mining applications.

Plant parts other than the grain of corn are also used in industry, forexample, stalks and husks are made into paper and wallboard and cobs areused for fuel and to make charcoal.

The seed of inbred corn line MN5, the plant produced from the inbredseed, the hybrid corn plant produced from the crossing of the inbred,hybrid seed, and various parts of the hybrid corn plant and transgenicversions of the foregoing, can be utilized for human food, livestockfeed, and as a raw material in industry.

Tables

In the tables that follow, the traits and characteristics of hybridcombinations having inbred corn line MN5 as a parental line are givencompared to other hybrids. The data collected are presented for keycharacteristics and traits. The field tests have been made at numerouslocations, with one or two replications per location. Information aboutthese hybrids as compared to the check hybrids is presented.

There may be a pedigree listed in the comparison group of the hybrid(s)containing an inbred parent with the nomenclature MON810 following theinbred name. MON810 is the designation given by the Monsanto Company(St. Louis, Mo.) for the transgenic event that, when expressed in maize,produces an endotoxin that is efficacious against the European cornborer, Ostrinia nubilalis, and certain other Lepidopteran larvae.Information for each pedigree includes:

1. Mean yield of the hybrid across all locations (bushels/acre) is shownin column 2.

2. A mean for the percentage moisture (H2O Grain) for the hybrid acrossall locations is shown in column 3.

3. A mean of the yield divided by the percentage moisture (Y/M) for thehybrid across all locations is shown in column 4.

4. A mean of the percentage of plants with stalk lodging (SL%) acrossall locations is shown in column 5.

5. A mean of the percentage of plants with root lodging (RL%) across alllocations is shown in column 6.

6. Test weight (TW) is the grain density as measured in pounds perbushel and is shown in column 7.

7. Harvest Appearance (Har Ap) is a rating made by a trained person onthe date of harvest. Harvest appearance is the rater's impression of thehybrid based on, but not limited to, a combination of factors to includeplant intactness, tissue health appearance and ease of harvest as itrelates to stalk lodging and root lodging. A scale of 1=Lowest to9=Highest/Most Desirable is used and is listed in column 8.

8. Plant Height (PlHt) is a physical measurement taken from the groundlevel to the tip of the tassel. It is expressed to the nearest inch andis shown in column 9.

9. Ear Height (EHt) is a physical measurement taken from the groundlevel to the node of attachment for the upper ear. It is expressed tothe nearest inch and is shown in column 10.

TABLE 2 Overall Comparisons: 2005/20 Tests Bushels/ H2O TW Har PlHt EHtPedigree Acre Grain Y/M SL % RL % lbs. Ap inches inches CB1 × MN5 207.420.0 10.4 2.4 8.7 55.4 6.2 113.5 42.0 KW4773 × MN5 200.3 20.0 10.0 1.64.6 55.7 6.2 112.9 43.0 CC2 × MN5 196.2 18.1 10.9 1.1 9.9 56.9 5.7 117.848.2 P33P67 201.8 20.1 10.0 6.4 3.0 56.6 5.8 115.8 44.9 P33N09 186.019.8 9.4 2.7 13.1 56.4 5.4 115.2 45.3 P34H31 170.4 18.8 9.1 1.3 7.6 57.15.2 111.9 41.7

TABLE 3 Overall Comparisons: 2005/18 Tests Bushels/ H2O TW Har PlHt EHtPedigree Acre Grain Y/M SL % RL % lbs. Ap inches inches CB1 × MN5 198.019.7 10.1 1.6 2.6 55.4 5.9 112.9 42.5 KW4773 × MN5 196.5 19.7 10.0 0.79.1 55.6 5.9 111.7 42.1 P33N09 187.0 20.2 9.2 5.3 9.0 55.9 5.7 113.340.9 P33D31 180.9 19.8 9.1 0.5 0.2 55.8 6.0 108.6 37.0

TABLE 4 Overall Comparisons: 2005/20 Tests Bushels/ H2O TW Har PlHt EHtPedigree Acre Grain Y/M SL % RL % lbs. Ap inches inches CC2 × MN5 194.617.7 11.0 5.9 3.3 56.8 5.8 123.3 48.0 LH331 × LH324 189.7 17.9 10.6 3.310.3 57.1 6.1 108.3 45.2 HC33 × LH326 181.3 16.7 10.9 2.8 6.3 57.7 4.8106.8 48.7 P33N09 187.9 19.7 9.5 4.6 12.0 56.5 5.6 117.0 44.8

TABLE 5 Overall Comparisons: 2005/20 Tests Bushels/ H2O TW Har PlHt EHtPedigree Acre Grain Y/M SL % RL % lbs. Ap inches inches BB14 × MN5 194.618.4 10.6 1.2 0.0 57.5 5.9 106.3 38.1 RBO1 × MN5 185.5 17.8 10.4 1.0 0.058.2 6.2 112.2 34.8 P34H31 179.8 18.1 9.9 1.2 0.1 58.3 5.5 104.3 34.8

TABLE 6 Overall Comparisons: 2004/10 Tests Bushels/ H2O TW Har PlHt EHtPedigree Acre Grain Y/M SL % RL % lbs. Ap inches inches RBO1 × MN5 210.819.2 11.4 0.8 0.0 57.5 6.3 111 39 RBO1 × LH287 207.6 17.8 12.0 0.7 0.057.8 6.0 114 35 HC33-ALS2 × LH287 206.4 18.8 11.3 0.1 0.1 57.5 6.3 11337 RBO1 × LH287Mon810 202.2 18.1 11.4 0.1 0.0 57.8 6.3 110 38

TABLE 7 Overall Comparisons: 2005/18 Tests Bushels/ H2O TW Har PlHt EHtPedigree Acre Grain Y/M SL % RL % lbs. Ap inches inches KW4773 × MN5184.2 19.5 9.4 2.1 8.3 55.5 5.7 116.8 44.2 LH311 × LH287Mon810 176.521.5 8.2 2.6 14.6 54.3 5.5 112.8 36.7 P33N09 169.5 19.9 8.5 10.2 7.856.1 5.2 118.1 42.3 LH310 × LH353 161.8 19.4 8.3 4.0 6.9 55.9 5.0 117.847.8

TABLE 8 Overall Comparisons: 2004/14 Tests Bushels/ H2O TW Har PlHt EHtPedigree Acre Grain Y/M SL % RL % lbs. Ap inches inches CB1 × MN5 218.817.2 12.9 0.6 0.0 57.2 6.8 115 47 LH311 × LH287Mon810 216.0 18.1 12.20.9 2.1 56.8 7.0 118 39 P33P67 211.5 16.7 12.9 1.3 0.6 59.3 6.4 117 42KW4773 × LH287Mon810 210.6 16.5 13.0 0.9 1.8 56.6 6.9 114 37

TABLE 9 Overall Comparisons: 2004/11 Tests Bushels/ H2O TW Har PlHt EHtPedigree Acre Grain Y/M SL % RL % lbs. Ap inches inches KW4773 × MN5219.8 20.2 11.3 0.1 1.2 56.2 6.9 122 46 P33P67 220.1 19.5 11.7 0.8 1.358.4 6.8 124 48 KW4635 × KW7638 218.5 19.9 11.3 0.8 1.2 56.9 6.7 117 42KWU4110 × KWU7104 217.6 18.3 12.3 0.7 0.6 57.3 6.7 112 48

TABLE 10 Overall Comparisons: 2003/6 Tests Bushels/ H2O TW Har PlHt EHtPedigree Acre Grain Y/M SL % RL % lbs. Ap inches inches KW4773 × MN5233.2 22.0 10.6 9.1 0.0 53.9 6.2 110 41 P33P67 232.7 22.0 10.9 8.3 0.054.6 7.2 108 37 HC33 × LH287 220.9 17.6 12.6 12.2 0.0 55.6 5.6 102 35RBO1 × LH287 211.6 17.3 12.3 10.1 2.1 55.6 5.0 102 35 KW4773 × LH287208.6 19.5 10.7 16.5 3.6 54.5 5.2 108 39

Deposit Information

Deposits of the inbred corn seed of this invention and the hybrid cornseed of this invention are maintained by AgReliant Genetics, 972 CountyRoad 500 East, Ivesdale, Ill. 61851. Access to these deposits will beavailable during the pendency of this application to persons determinedby the Commissioner of Patents and Trademarks to be entitled theretounder 37 C.F.R. §1.14 and 35 U.S.C. §122. Upon allowance of one or moreclaims directed to the inbred corn seed of this invention, allrestrictions on the availability to the public of the inbred will beirrevocably removed by affording access to a deposit of at least 2,500seeds of the same inbred with the National Collections of Industrial,Food and Marine Bacteria (NCIMB), 23 St Machar Drive, Aberdeen,Scotland, AB24 3RY, United Kingdom. Upon allowance of one or more claimsdirected to one or more hybrids of this invention, all restrictions onthe availability to the public of the hybrid(s) will be irrevocablyremoved by affording access to a deposit of at least 2,500 seeds of thesame hybrid(s) with the National Collections of Industrial, Food andMarine Bacteria (NCIMB), 23 St Machar Drive, Aberdeen, Scotland, AB243RY, United Kingdom.

While a number of exemplary aspects and embodiments have been discussedabove, those of skill in the art will recognize certain modifications,permutations, additions and sub-combinations thereof. It is thereforeintended that the following appended claims and claims hereafterintroduced are interpreted to include all such modifications,permutations, additions and sub-combinations as are within their truespirit and scope.

1. A seed of corn inbred line designated MN5, wherein a representativesample of seed of said line was deposited under NCIMB ______.
 2. A cornplant, or a part thereof, produced by growing the seed of claim
 1. 3. Acorn plant, or a part thereof, having all the physiological andmorphological characteristics of the inbred line MN5, wherein arepresentative sample of seed of said line was deposited under NCIMB______.
 4. A tissue culture of cells produced from the plant of claim 2,wherein said cells of the tissue culture are produced from a plant partselected from the group consisting of leaves, pollen, embryos, roots,root tips, anthers, silks, flowers, kernels, ears, cobs, husks, seedsand stalks.
 5. A corn plant regenerated from the tissue culture of claim4, wherein the regenerated plant has all the morphological andphysiological characteristics of inbred line MN5, wherein arepresentative sample of seed of said line was deposited under NCIMB______.
 6. A method for producing a hybrid corn seed wherein the methodcomprises crossing the plant of claim 2 with a different corn plant andharvesting the resultant hybrid corn seed.
 7. A hybrid corn seedproduced by the method of claim
 6. 8. A method for producing inbred lineMN5, wherein a representative sample of seed of said line was depositedunder NCIMB ______, wherein the method comprises: a) planting acollection of seed comprising seed of a hybrid, one of whose parents isinbred line MN5, said collection also comprising seed of said inbred; b)growing plants from said collection of seed; c) identifying the plantshaving the physiological and morphological characteristics of corninbred line MN5 as inbred parent plants; d) controlling pollination ofsaid inbred parent plants in a manner which preserves the homozygosityof said inbred parent plant; and e) harvesting the resultant seed. 9.The method of claim 8 wherein step (c) comprises identifying plants withdecreased vigor compared to the other plants grown from the collectionof seeds.
 10. A method for producing a corn plant that contains in itsgenetic material one or more transgenes, wherein the method comprisescrossing the corn plant of claim 2 with either a second plant of anothercorn line which contains a transgene or a transformed corn plant of theinbred corn line MN5, so that the genetic material of the progeny thatresults from the cross contains the transgene(s) operably linked to aregulatory element and wherein the transgene is selected from the groupconsisting of male sterility, male fertility, herbicide resistance,insect resistance, disease resistance, water stress tolerance, andincreased digestibility.
 11. A corn plant, or a part thereof, producedby the method of claim
 10. 12. The corn plant of claim 11, wherein thetransgene confers resistance to an herbicide selected from the groupconsisting of imidazolinone, sulfonylurea, glyphosate, glufosinate,L-phosphinothricin, triazine and benzonitrile.
 13. The corn plant ofclaim 11, wherein the transgene encodes a Bacillus thuringiensisprotein.
 14. The corn plant of claim 11, wherein the transgene confersdisease resistance.
 15. The corn plant of claim 11, wherein thetransgene confers water stress tolerance.
 16. The corn plant of claim11, where in the transgene confers increased digestibility.
 17. A methodfor producing a hybrid corn seed wherein the method comprises crossingthe plant of claim 11 with a different corn plant and harvesting theresultant hybrid corn seed.
 18. A method of producing a corn plant withwaxy starch or increased amylose starch wherein the method comprisestransforming the corn plant of claim 2 with a transgene that modifiescarbohydrate metabolism.
 19. A corn plant produced by the method ofclaim
 18. 20. A method of introducing a desired trait into corn inbredline MN5 wherein the method comprises: a) crossing the inbred line MN5plants grown from the inbred line MN5 seed, wherein a representativesample of seed of said line was deposited under NCIMB ______, withplants of another corn line that comprise a desired trait to produceprogeny plants, wherein the desired trait is selected from the groupconsisting of male sterility, male fertility, herbicide resistance,insect resistance, disease resistance, waxy starch, water stresstolerance, increased amylose starch and increased digestibility; b)selecting progeny plants that have the desired trait to produce selectedprogeny plants; c) crossing the selected progeny plants with the inbredline MN5 plants to produce backcross progeny plants; d) selecting forbackcross progeny plants that have the desired trait and physiologicaland morphological characteristics of corn inbred line MN5 listed inTable 1 to produce selected backcross progeny plants; and e) repeatingsteps (c) and (d) one or more times in succession to produce selectedsecond or higher backcross progeny plants that comprise the desiredtrait and the physiological and morphological characteristics of corninbred line MN5 as listed in Table
 1. 21. A corn plant produced by themethod of claim 20, wherein the plant has the desired trait and all ofthe physiological and morphological characteristics of corn inbred lineMN5 as listed in Table
 1. 22. A method for producing inbred line MN5seed, wherein a representative sample of seed of said line was depositedunder NCIMB ______, wherein the method comprises crossing a first inbredparent corn plant with a second inbred parent corn plant and harvestingthe resultant corn seed, wherein both said first and second inbred cornplant are the corn plant of claim
 3. 23. A method for producing inbredline MN5 seed, wherein a representative sample of seed of said line wasdeposited under NCIMB ______, wherein the method comprises: a) plantingan inbred corn seed of claim 1; b) growing plant from said seed; c)controlling pollination in a manner that the pollen produced by thegrown plant pollinates the ovules produced by the grown plant; and d)harvesting the resultant seed.
 24. Hybrid corn seed designated CB1*MN5having inbred line MN5 or a transgenic inbred line of MN5 as a parentalline, wherein a representative sample of seed of MN5 was deposited underNCIMB ______, and having inbred line CB1 or a transgenic inbred line ofCB1 as a parental line, wherein a representative sample of seed of CB1was deposited under NCIMB No. 41375, wherein the transgenic inbred lineMN5 or CB1 comprises a transgene that confers a trait selected from thegroup consisting of disease resistance, male sterility, herbicideresistance, and insect resistance.
 25. Hybrid corn seed designatedKW4773*MN5 having inbred line MN5 or a transgenic inbred line of MN5 asa parental line, wherein a representative sample of seed of MN5 wasdeposited under NCIMB ______, and having inbred line KW4773 or atransgenic inbred line of KW4773 as a parental line, wherein arepresentative sample of seed of KW47731 was deposited under ATCCAccession No. PTA-5307, wherein the transgenic inbred line MN5 or KW4773comprises a transgene that confers a trait selected from the groupconsisting of disease resistance, male sterility, herbicide resistance,and insect resistance.
 26. Hybrid corn seed designated CC2*MN5 havinginbred line MN5 or a transgenic inbred line of MN5 as a parental line,wherein a representative sample of seed of MN5 was deposited under NCIMB______, and having inbred line CC2 or a transgenic inbred line of CC2 asa parental line, wherein a representative sample of seed of CC2 wasdeposited under NCIMB No. 41376, wherein the transgenic inbred line MN5or CC2 comprises a transgene that confers a trait selected from thegroup consisting of disease resistance, male sterility, herbicideresistance, and insect resistance.
 27. Hybrid corn seed designatedBB14*MN5 having inbred line MN5 or a transgenic inbred line of MN5 as aparental line, wherein a representative sample of seed of MN5 wasdeposited under NCIMB ______, and having inbred line BB14 or atransgenic inbred line of BB14 as a parental line, wherein arepresentative sample of seed of BB14 was deposited under NCIMB ______,wherein the transgenic inbred line MN5 or BB14 comprises a transgenethat confers a trait selected from the group consisting of diseaseresistance, male sterility, herbicide resistance, and insect resistance.28. Hybrid corn seed designated RBO1*MN5 having inbred line MN5 or atransgenic inbred line of MN5 as a parental line, wherein arepresentative sample of seed of MN5 was deposited under NCIMB ______,and having inbred line RBO1 or a transgenic inbred line of RBO1 as aparental line, wherein a representative sample of seed of RBO1 wasdeposited under ATCC Accession No. PTA-5627, wherein the transgenicinbred line MN5 or RBO1 comprises a transgene that confers a traitselected from the group consisting of disease resistance, malesterility, herbicide resistance, and insect resistance.